Decreasing the postprandial glucose response is potentially of major importance to public health when low-glycemic index or high-fibre content foods are associated with a decreased risk of diabetes. We investigated in overweight subjects the effect of adding beta-glucan (BG) to a polenta (Pol) meal on postprandial metabolism and glucose bioavailability using stable isotopes. In this single-blind, randomized, crossover trial, 12 subjects ate two meals containing Pol with (Pol + BG) or without (Pol) 5 g BG. Concentrations of glucose, insulin, C-peptide, nonesterified fatty acids, triacylglycerol, total and exogenous glucose kinetics were assessed for 6 h postprandially. The kinetics of total and exogenous glucose importantly differed between the meals, but not the quantity of total and exogenous glucose appearing in plasma. Less total and exogenous glucose appeared during the first 120 min after the Pol + BG meal; the phenomenon was then reversed (both p < 0.0001). After 120 min, glucose and insulin responses declined, but remained higher after the Pol + BG meal (p < 0.05) in parallel to the inhibition of lipolysis. The endogenous glucose production (EGP) was significantly more inhibited after the Pol + BG meal. The addition of BG slowed the appearance of glucose in plasma, resulting in longer-lasting insulin secretion which exerted a prolonged inhibition of EGP and lipolysis.